CD45 is a receptor-like protein tyrosine phosphatase highly expressed on all nucleated hematopoietic cells. We previously generated mice containing a point mutation in the juxtamembrane wedge of CD45. Demonstrating the critical negative regulatory function of the wedge, the CD45 E613R mutation led to a lymphoproliferative disorder (LPD) and a lupus-like autoimmune syndrome. Here we show the central role of B cells in this phenotype. Genetic elimination of B cells, but not T cells, ablates the LPD. In contrast to CD45-deficient B cells, the E613R mutation generates hyperresponsive B cells. Comparison of CD45-deficient and CD45 E613R mice reveals dichotomous effects of these mutations on B cell development. Together, the results support a role for CD45 as a rheostat, with both positive and negative regulatory functions, that fine-tunes the signal transduction threshold at multiple checkpoints in B cell development.

The cooperative activity of protein tyrosine kinases and phosphatases plays a central role in regulation of T cell receptor (TCR) signal strength. Perturbing this balance, and thus the threshold for TCR signals, has profound impacts on T cell development and function. We previously generated mice containing a point mutation in the juxtamembrane wedge of the receptor-like protein tyrosine phosphatase CD45. Demonstrating the critical negative regulatory function of the wedge, the CD45 E613R (WEDGE) mutation led to a lymphoproliferative disorder (LPD) and a lupus-like autoimmune syndrome. Using genetic, cellular, and biochemical approaches, we now demonstrate that the CD45 wedge influences T cell development and function. Consistent with increased TCR signal strength, WEDGE mice have augmented positive selection and enhanced sensitivity to the CD4-mediated disease experimental autoimmune encephalitis (EAE). These correspond with hyperresponsive calcium and pERK responses to TCR stimulation in thymocytes, but surprisingly, not in peripheral T cells, where these responses are actually depressed. Together, the data support a role for the CD45 wedge in regulation of T cell responses in vivo and suggest that its effects depend on cellular context. autoimmunity ͉ tyrosine phosphatase ͉ tyrosine kinase ͉ thymocyte development T cell receptor (TCR) signal strength is influenced by the integration of multiple inputs including affinity for antigen, presence and activity of costimulatory molecules, and duration of the interaction with antigen-presenting cells (APCs) (1). Alterations in TCR signal strength impact many aspects of T cell biology including CD4 versus CD8 lineage commitment (1), effector and memory cell generation (2), and autoimmunity (3, 4). Currently, the factors defining signal strength and its functional outcome are incompletely understood.CD45, a receptor-like protein tyrosine phosphatase (RPTP) expressed on all nucleated hematopoietic cells, plays a critical positive regulatory role in antigen receptor signaling. Its absence in both mice and humans results in severe combined immunodeficiency (5, 6). CD45 mediates its effects, at least in part, by modulating the activation state of Src family protein tyrosine kinases (SFKs) (5, 7). Phosphorylation of the SFK C-terminal tyrosine by Csk inhibits the kinase while autophosphorylation of the catalytic domain tyrosine results in full activity. CD45 opposes Csk by dephosphorylating the negative regulatory tyrosine, generating a pool of primed SFKs capable of rapid activation upon receptor stimulation. In some cell types, CD45 can also dephosphorylate the catalytic tyrosine and negatively regulate SFKs.Regulation of CD45 itself is complex. Alternative splicing of the extracellular domain, regulated in a cell-and activation-specific manner, generates multiple isoforms differing in size and charge (5). Interestingly, CD45 polymorphisms influencing its alternative splicing, and thus isoform expression, are associated with several human autoimmune diseases (8). We initi...

The autoimmune disease systemic lupus erythematosus (SLE) has a complex environmental and multi-factorial genetic basis. Genome wide association studies have recently identified numerous disease-associated polymorphisms, but it remains unclear in which cells and during which step of pathogenesis specific polymorphisms interact to cause disease. Using a mouse model in which the same activating mutation (CD45E613R) causes distinct genetic background-dependent disease phenotypes, we perform a screen for genetic modifiers of autoreactivity between anti-nuclear antibody (ANA)-resistant CD45E613R.B6 and ANA-permissive CD45E613R.BALB/c mice. Within a novel autoreactivity-associated locus on chromosome 9, we identify a putative modifier, TLR9. Validating a role for TLR9 in modifying autoreactivity in the context of the CD45E613R mutation, manipulation of TLR9 gene dosage eliminates ANA in CD45E613R.BALB/c, but confoundingly permits ANA in CD45E613R.B6. We demonstrate that sensitivity to ANA is modulated by strength of TLR9 signal, since stronger TLR9B6 signals, but not weaker TLR9BALB/c signals, negatively regulate CD45E613R B cell development during competitive reconstitution at the central tolerance checkpoint. Our results identify a novel autoreactivity-associated locus and validate Tlr9 as a candidate gene within the locus. We further demonstrate a novel role for TLR9 signal strength in central tolerance, providing insight into the interplay of disease-associated polymorphisms at a discrete step of SLE pathogenesis.